c-Jun N-terminal Kinase Signaling Research Articles

Protective Role of microRNA-31 in Acetaminophen-Induced Liver Injury: A Negative Regulator of c-Jun N-Terminal Kinase (JNK) Signaling Pathway

Background & AimsSustained c-Jun N-terminal kinase (JNK) activation plays a major role in drug-induced liver injury (DILI). Stress-responsive microRNA-31 (miR-31) has been implicated in regulating different cellular damage, and JNK activation could induce miR-31 expression. However, the regulatory role of miR-31 in DILI has not been studied previously. We aimed to investigate whether miR-31 could ameliorate DILI and ascertain potential molecular mechanism.MethodsmiR-31 gene knockout (31-KO) and wild-type C57BL/6J mice were used to construct an acetaminophen (APAP)-induced DILI model. Primary mouse hepatocytes, as well as alpha mouse liver 12 (AML-12) cell lines, were used for in vitro experiments. Argonaute 2–associated RNA immunoprecipitation combined with high-throughput sequencing were performed to identify specific targets of miR-31.Results31-KO mice showed a higher mortality rate, liver transaminase levels, and hepatic necrosis compared with those in wild-type mice after APAP-induced hepatotoxicity. The protective role of miR-31 on hepatocytes has been analyzed via constructing bone marrow chimeric mice. Mechanistically, we found that hepatic JNK phosphorylation increased significantly in 31-KO mice. This caused mitochondrial phosphorylated Src (p-Src) inactivation and more reactive oxygen species production, which directly amplifies hepatocyte necrotic cell death, while administration of JNK-specific inhibitor SP600125 could abrogate the differences. Moreover, bioinformatics analysis of RNA immunoprecipitation combined with high-throughput sequencing identified that guanosine triphosphatase, cell division cycle protein 42 (Cdc42), the upstream molecule of JNK signaling, was the specific target of miR-31 and could form a miR-31/Cdc42/phosphorylated mixed-lineage kinase 3 (p-MLK3) negative feedback loop to restrict JNK overactivation. Clinically, both miR-31 and phosphorylated JNK (p-JNK) were highly increased in liver tissues of DILI patients with different etiologies.ConclusionsmiR-31 can down-regulate Cdc42 to restrict overactivation of reactive oxygen species/JNK/mitochondria necrotic death loop in hepatocytes of APAP-induced DILI, which might provide a new therapeutic target for alleviating JNK overactivation–based liver injury.

The Anti-inflammatory Effects of the Bioactive Compounds Isolated from Alpinia officinarum Hance Mediated by the Suppression of NF-kappaB and MAPK Signaling.

This study was designed to evaluate the anti-inflammatory effects of Alpinia officinarum Hance extract (AOE) and identify its main active ingredients. AOE was obtained using a 95% ethanol extraction method. Lipopolysaccharide (LPS) were used to induce an inflammatory response in RAW264.7 cells. The results showed that AOE exerts anti-inflammatory effects via inhibition of prostaglandin E2 secretion and cyclooxygenase -2 (COX-2) production. We further analyzed the components of AOE using high-performance liquid chromatography and found that AOE is comprised of several bioactive flavonoids including quercetin (Q), kaempferol (K), galangin (G), and curcumin (C). These four flavonoids effectively inhibited nitric oxide (NO), interleukin (IL)-1β, IL-6, and tumor necrosis factor-α production. Moreover, they reduced COX-2 and inducible NO synthase expressions via regulation of nuclear factor kappa-light-chain-enhancer of activated B cells and c-Jun N-terminal kinase signaling pathways. Furthermore, we compared and contrasted the anti-inflammatory effects and mechanisms of these four flavonoids at the same dose in the LPS-induced cell inflammation model. The results showed that C is the most effective inhibitor of LPS-induced NO production. However, only Q and K effectively attenuated LPS-induced extracellular signal-regulated kinase and p38 elevations. In conclusion, AOE and its major bioactive compounds exert anti-inflammatory effects on LPS-induced inflammation. As A. officinarum Hance is much cheaper than any of its four flavonoids, especially G, we suggest using AOE as an anti-inflammatory agent.

HSA_CIRC_0004050 on proliferation and apoptosis of A549 cells through ERK/JNK signaling pathway.

The aim of this study was to screen the differentially expressed circular ribonucleic acid (circRNA) in non-small cell lung cancer (NSCLC) and explore its functional mechanism. Differentially expressed circRNAs in tumor tissues of NSCLC patients were detected via gene microarray and reverse transcriptionquantitative polymerase chain reaction (RT-qPCR). The associaton between their expressions and the clinical phenotypes was explored combined with clinical data. The effect of overexpression of hsa_circ_0004050 on the proliferation of A549 cells was detected via cell counting kit-8 (CCK-8) assay and CFSE assay. The effect of overexpression of hsa_circ_0004050 (human circular ribonucleic acid_0004050) on the apoptosis of A549 cells was detected using the Annexin V-FITC/PI kit. Then the direct-acting miRNAs of hsa_circ_0004050 were screened using bioinformatics software and luciferase reporter assay, and the direct targets of miR- 1233-3p were explored using bioinformatics software and luciferase reporter assay combined with RTqPCR and Western blotting. The effects of overexpression of miR-1233-3p or knockdown of dual specificity phosphatase 9 (DUSP9) on the cell proliferation and apoptosis affected by overexpression of hsa_circ_0004050 were detected. Western blotting was performed to detect the effects of hsa_circ_0004050 on the extracellular signal-regulated kinase (ERK)/c-Jun N-terminal kinase (JNK) signaling pathway. The expression of hsa_ circ_0004050 was significantly lower in tumor tissues than that in para-carcinoma tissues in NSCLC patients. The expression of hsa_circ_0004050 was significantly correlated with TNM stage, tumor size and lymph node metastasis. The results of survival analysis showed that the survival time of patients with a high expression of hsa_circ_0004050 was obviously prolonged. According to the results of phenotype assay, hsa_circ_0004050 could promote apoptosis and inhibit proliferation of A549 cells. In terms of its mechanism, hsa_circ_0004050 could markedly increase the protein expression of DUSP9 via targeting miR-1233-3p in A549 cells, thereby inhibiting the ERK/JNK signaling pathway. Hsa_circ_0004050 may serve as a potential therapeutic target for NSCLC or a biomarker for the diagnosis of NSCLC in the future.

E6-mediated activation of JNK drives EGFR signalling to promote proliferation and viral oncoprotein expression in cervical cancer

Human papillomaviruses (HPV) are a major cause of malignancy worldwide, contributing to ~5% of all human cancers including almost all cases of cervical cancer and a growing number of ano-genital and oral cancers. HPV-induced malignancy is primarily driven by the viral oncogenes, E6 and E7, which manipulate host cellular pathways to increase cell proliferation and enhance cell survival, ultimately predisposing infected cells to malignant transformation. Consequently, a more detailed understanding of viral-host interactions in HPV-associated disease offers the potential to identify novel therapeutic targets. Here, we identify that the c-Jun N-terminal kinase (JNK) signalling pathway is activated in cervical disease and in cervical cancer. The HPV E6 oncogene induces JNK1/2 phosphorylation in a manner that requires the E6 PDZ binding motif. We show that blockade of JNK1/2 signalling using small molecule inhibitors, or knockdown of the canonical JNK substrate c-Jun, reduces cell proliferation and induces apoptosis in cervical cancer cells. We further demonstrate that this phenotype is at least partially driven by JNK-dependent activation of EGFR signalling via increased expression of EGFR and the EGFR ligands EGF and HB-EGF. JNK/c-Jun signalling promoted the invasive potential of cervical cancer cells and was required for the expression of the epithelial to mesenchymal transition (EMT)-associated transcription factor Slug and the mesenchymal marker Vimentin. Furthermore, JNK/c-Jun signalling is required for the constitutive expression of HPV E6 and E7, which are essential for cervical cancer cell growth and survival. Together, these data demonstrate a positive feedback loop between the EGFR signalling pathway and HPV E6/E7 expression, identifying a regulatory mechanism in which HPV drives EGFR signalling to promote proliferation, survival and EMT. Thus, our study has identified a novel therapeutic target that may be beneficial for the treatment of cervical cancer.

Isoliquiritigenin Enhances the Beige Adipocyte Potential of Adipose-Derived Stem Cells by JNK Inhibition.

Human adipose-derived stem cells (hASCs) can be isolated from fat tissue and have attracted interest for their potential therapeutic applications in metabolic disease. hASCs can be induced to undergo adipogenic differentiation in vitro by exposure to chemical agents or inductive growth factors. We investigated the effects and mechanism of differentiating hASC-derived white adipocytes into functional beige and brown adipocytes with isoliquiritigenin (ILG) treatment. Here, we showed that hASC-derived white adipocytes could promote brown adipogenesis by expressing both uncoupling protein 1 (UCP1) and PR/SET Domain 16 (PRDM16) following low-dose ILG treatments. ILG treatment of white adipocytes enhanced the expression of brown fat-specific markers, while the expression levels of c-Jun N-terminal kinase (JNK) signaling pathway proteins were downregulated. Furthermore, we showed that the inhibition of JNK phosphorylation contributed to white adipocyte differentiation into beige adipocytes, which was validated by the use of SP600125. We identified distinct regulatory effects of ILG dose responses and suggested that low-dose ILG induced the beige adipocyte potential of hASCs via JNK inhibition.

Oral Administration of the p75 Neurotrophin Receptor Modulator, LM11A-31, Improves Erectile Function in a Mouse Model of Cavernous Nerve Injury

BackgroundRadical prostatectomy for prostate cancer can not only induce cavernous nerve injury (CNI), but also causes cavernous hypoxia and cavernous structural changes, which lead to a poor response to phosphodiesterase 5 inhibitors. AimTo investigate the therapeutic effect of oral administration of LM11A-31, a small molecule p75 neurotrophin receptor (p75NTR) ligand and proNGF antagonist, in a mouse model of bilateral CNI, which mimics nerve injury–induced erectile dysfunction after radical prostatectomy. Methods8-week-old male C57BL/6 mice were divided into sham operation and CNI groups. Each group was divided into 2 subgroups: phosphate-buffered saline and LM11A-31 (50 mg/kg/day) being administered once daily starting 3 days before CNI via oral gavage. 2 weeks after CNI, we measured erectile function by electrical stimulation of the bilateral cavernous nerve. The penis was harvested for histologic examination and Western blot analysis. The major pelvic ganglia was harvested and cultured for assays of ex vivo neurite outgrowth. OutcomesIntracavernous pressure, neurovascular regeneration in the penis, in vivo or ex vivo functional evaluation, and cell survival signaling were measured. ResultsErectile function was decreased in the CNI group (44% of the sham operation group), while administration of LM11A-31 led to a significant improvement of erectile function (70% of the sham operation group) in association with increased neurovascular content, including cavernous endothelial cells, pericytes, and neuronal processes. Immunohistochemical and Western blot analyses showed significantly increased p75NTR expression in the dorsal nerve of CNI mice, which was attenuated by LM11A-31 treatment. Protein expression of active PI3K, AKT, and endothelial nitric oxide synthase was increased, and cell death and c-Jun N-terminal kinase signaling was significantly attenuated after LM11A-31 treatment. Furthermore, LM11A-31 promoted neurite sprouting in cultured major pelvic ganglia after lipopolysaccharide exposure. Clinical ImplicationsLM11A-31 may be used as a strategy to treat erectile dysfunction after radical prostatectomy or in men with neurovascular diseases. Strengths & LimitationsUnlike biological therapeutics, such as proteins, gene therapies, or stem cells, the clinical application of LM11A-31 would likely be relatively less complex and low cost. Our study has some limitations. Future studies will assess the optimal dosing and duration of the compound. Given its plasma half-life of approximately 1 hour, it is possible that dosing more than once per day will provide added efficacy. ConclusionSpecific inhibition of the proNGF-p75NTR degenerative signaling via oral administration of LM11A-31 represents a novel therapeutic strategy for erectile dysfunction induced by nerve injury.Yin GN, Ock J, Limanjaya A, et al. Oral Administration of the p75 Neurotrophin Receptor Modulator, LM11A-31, Improves Erectile Function in a Mouse Model of Cavernous Nerve Injury. J Sex Med 2021;18:17–28.

Acacetin Induces Apoptosis in Human Osteosarcoma Cells by Modulation of ROS/JNK Activation.

PurposeThe long-term survival rate of osteosarcoma, which is the most common type of primary malignant bone tumor, has stagnated in past decades. Acacetin is a natural flavonoid compound that has antioxidative and anti-inflammatory effects and exhibits extensive therapeutic effects on various cancers. In this study, the anticancer potential of acacetin and the underlying molecular mechanisms were examined in human osteosarcoma cells (SJSA and HOS).Materials and MethodsHOS and SJSA cell lines were exposed to different concentrations of acacetin. Cell proliferation and viability were assessed by CCK-8 and colony-formation assays. Hoechst 33258 fluorescent staining was employed to detect apoptosis. Cell apoptosis was measured by an annexin V-FITC/PI assay by flow cytometry. The alteration in the mitochondrial membrane potential was detected by a JC-1 Assay Kit. Apoptosis-related protein expression was determined by Western blotting. Intracellular reactive oxygen species (ROS) production was detected by fluorescence microscopy and flow cytometry. Subsequently, the activation of the ROS/JNK signaling pathway was investigated.ResultsAcacetin could inhibit proliferation and induce apoptosis in SJSA and HOS cells. The acacetin treatment resulted in the activation of caspase-3, −8, and −9 and cleaved PARP. Further studies showed that acacetin-induced apoptosis was attributed to ROS. In addition, we found that acacetin induced the activation of the downstream c-Jun N-terminal kinase (JNK) signaling pathway. Subsequently, after treatment with the ROS scavenger GSH and the JNK inhibitor SP600125, the apoptosis-inducing effect triggered by acacetin was significantly attenuated.ConclusionThe results of the present study indicate that acacetin may induce apoptosis to inhibit cell growth by activating the ROS/JNK signaling pathway in SJSA and HOS cells, suggesting that acacetin may be a promising candidate for the management of osteosarcomas.

Protective Role of St. John's Wort and Its Components Hyperforin and Hypericin against Diabetes through Inhibition of Inflammatory Signaling: Evidence from In Vitro and In Vivo Studies.

Diabetes mellitus is a very common chronic disease with progressively increasing prevalence. Besides the well-known autoimmune and inflammatory pathogenesis of type 1 diabetes, in many people, metabolic changes and inappropriate lifestyle favor a subtle chronic inflammatory state that contributes to development of insulin resistance and progressive loss of β-cell function and mass, eventually resulting in metabolic syndrome or overt type 2 diabetes. In this paper, we review the anti-inflammatory effects of the extract of Hypericum perforatum L. (St. John’s wort, SJW) and its main active ingredients firstly in representative pathological situations on inflammatory basis and then in pancreatic β cells and in obese or diabetic animal models. The simultaneous and long-lasting inhibition of signal transducer and activator of transcription (STAT)-1, nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) and mitogen-activated protein kinases (MAPKs)/c-jun N-terminal kinase (JNK) signaling pathways involved in pro-inflammatory cytokine-induced β-cell dysfunction/death and insulin resistance make SJW particularly suitable for both preventive and therapeutic use in metabolic diseases. Hindrance of inflammatory cytokine signaling is likely dependent on the hyperforin content of SJW extract, but recent data reveal that hypericin can also exert relevant protective effects, mediated by activation of the cyclic adenosine monophosphate (cAMP)/protein kinase cAMP-dependent (PKA)/adenosine monophosphate activated protein kinase (AMPK) pathway, against high-fat-diet-induced metabolic abnormalities. Actually, the mechanisms of action of the two main components of SJW appear complementary, strengthening the efficacy of the plant extract. Careful quantitative analysis of SJW components and suitable dosage, with monitoring of possible drug–drug interaction in a context of remarkable tolerability, are easily achievable pre-requisites for forthcoming clinical applications.

Hypothermia Advocates Functional Mitochondria and Alleviates Oxidative Stress to Combat Acetaminophen-Induced Hepatotoxicity.

For years, moderate hypothermia (32 °C) has been proposed as an unorthodox therapy for liver injuries, with proven hepatoprotective potential. Yet, limited mechanistic understanding has largely denied its acceptance over conventional pharmaceuticals for hepatoprotection. Today, facing a high prevalence of acetaminophen-induced liver injury (AILI) which accounts for the highest incidence of acute liver failure, hypothermia was evaluated as a potential therapy to combat AILI. For which, transforming growth factor-α transgenic mouse hepatocytes (TAMH) were subjected to concomitant 5 mM acetaminophen toxicity and moderate hypothermic conditioning for 24 h. Thereafter, its impact on mitophagy, mitochondrial biogenesis, glutathione homeostasis and c-Jun N-terminal kinase (JNK) signaling pathways were investigated. In the presence of AILI, hypothermia displayed simultaneous mitophagy and mitochondrial biogenesis to conserve functional mitochondria. Furthermore, antioxidant response was apparent with higher glutathione recycling and repressed JNK activation. These effects were, however, unremarkable with hypothermia alone without liver injury. This may suggest an adaptive response of hypothermia only to the injured sites, rendering it favorable as a potential targeted therapy. In fact, its cytoprotective effects were displayed in other DILI of similar pathology as acetaminophen i.e., valproate- and diclofenac-induced liver injury and this further corroborates the mechanistic findings of hypothermic actions on AILI.

Effect of pristimerin on apoptosis through activation of ROS/ endoplasmic reticulum (ER) stress-mediated noxa in colorectal cancer

BackgroundPristimerin, a natural quinonemethid triterpenoid found in different spp. of Celastraceae and Hippocrateaceae families, has been reported to exhibit potent antitumor activities against colorectal cancer (CRC). However, the mechanisms underlying pristimerin-induced apoptosis in CRC is not clear. PurposeThis study aimed to investigate the mechanisms of pristimerin-induced apoptosis against CRC in vitro and in vivo. MethodsCell viability and cell apoptosis analyses were conducted to assess the effects of pristimerin on CRC. Western blotting was performed to detect the expression of proteins affected by pristimerin in vitro and in vivo. HCT116 colon cancer xenografts and APCmin/+ mouse models were used to evaluate the anti-CRC effect of pristimerin in vivo. ResultsOur data showed that pristimerin induced apoptosis by regulating proapoptotic proteins of which Noxa showed higher expression. Pristimerin triggered reactive oxygen species (ROS)-mediated endoplasmic reticulum (ER) stress signaling activation. Pristimerin significantly elevated the expression of ER stress-related proteins, resulting in activation of the IRE1α and c-Jun N-terminal kinase (JNK) signal pathway through the formation of the IRE1α-TRAF2-ASK1 complex. Pristimerin exhibited apoptosis-inducing activities in HCT116 colon cancer xenografts and APCmin/+ mice. ConclusionBoth in vitro and in vivo data demonstrated that pristimerin induced Noxa expression and apoptosis through activation of the ROS/ER stress/JNK axis in CRC. Thus, pristimerin may be a promising antitumor agent for CRC.

Alarmins and c-Jun N-Terminal Kinase (JNK) Signaling in Neuroinflammation.

Neuroinflammation is involved in the progression or secondary injury of multiple brain conditions, including stroke and neurodegenerative diseases. Alarmins, also known as damage-associated molecular patterns, are released in the presence of neuroinflammation and in the acute phase of ischemia. Defensins, cathelicidin, high-mobility group box protein 1, S100 proteins, heat shock proteins, nucleic acids, histones, nucleosomes, and monosodium urate microcrystals are thought to be alarmins. They are released from damaged or dying cells and activate the innate immune system by interacting with pattern recognition receptors. Being principal sterile inflammation triggering agents, alarmins are considered biomarkers and therapeutic targets. They are recognized by host cells and prime the innate immune system toward cell death and distress. In stroke, alarmins act as mediators initiating the inflammatory response after the release from the cellular components of the infarct core and penumbra. Increased c-Jun N-terminal kinase (JNK) phosphorylation may be involved in the mechanism of stress-induced release of alarmins. Putative crosstalk between the alarmin-associated pathways and JNK signaling seems to be inherently interwoven. This review outlines the role of alarmins/JNK-signaling in cerebral neurovascular inflammation and summarizes the complex response of cells to alarmins. Emerging anti-JNK and anti-alarmin drug treatment strategies are discussed.

Prevention of Progression of Aortic Aneurysm by Peptide Vaccine Against Ang II (Angiotensin II) in a Rat Model

There is no proven medical therapy to inhibit the progression of abdominal aortic aneurysm (AAA) in the clinical setting. To develop a novel therapeutic approach for the treatment of AAA, we focused on vaccination targeting Ang II (angiotensin II) and assessed the effect of an Ang II peptide vaccine on the progression of AAA using a rat model. Ang II peptide was conjugated with KLH (keyhole limpet hemocyanin) carrier protein to induce a sufficient immune response. Male rats were subcutaneously immunized with Ang II-KLH with an adjuvant on days 0, 14, and 28. Aortic dilatation was induced by intraluminal incubation with elastase on day 35. Treatment with Ang II vaccine successfully induced the production of a high titer of anti-Ang II antibodies. Immunization with Ang II vaccine resulted in a significant reduction in expansion of the aortic diameter compared with control rats, without a blood pressure-lowering effect. Four weeks after operation, the increase in Ang II in the aneurysm wall was significantly inhibited by treatment with Ang II vaccine. Inhibition of Ang II action led to suppression of the inflammatory response in the AAA wall through attenuation of the NFκB (nuclear factor kappa B) and c-jun N-terminal kinase signaling cascades. Treatment with Ang II vaccine inhibited accumulation of macrophages in the AAA wall. In addition, expression of TNF-α (tumor necrosis factor alpha) and activation of MMP (matrix metalloproteinase)-2 and MMP-9 were also inhibited by treatment with Ang II vaccine, resulting in protection against the destruction of elastic fibers. This vaccine therapy could become a potent therapeutic option to treat patients with AAA.

IL-1/IL-1R signaling induced by all-trans-retinal contributes to complement alternative pathway activation in retinal pigment epithelium.

The underlying mechanisms of complement activation in Stargardt disease type 1 (STGD1) and age-related macular degeneration (AMD) are not fully understood. Overaccumulation of all-trans-retinal (atRAL) has been proposed as the pathogenic factor in both diseases. By incubating retinal pigment epithelium (RPE) cells with atRAL, we showed that C5b-9 membrane attack complexes (MACs) were generated mainly through complement alternative pathway. An increase in complement factor B (CFB) expression as well as downregulation of complement regulatory proteins CD46, CD55, CD59, and CFH were observed in RPE cells after atRAL treatment. Furthermore, interleukin-1β production was provoked in both atRAL-treated RPE cells and microglia/macrophages. Coincubation of RPE cells with interleukin-1 receptor antagonist (IL1Ra) and atRAL ameliorated complement activation and downregulated CFB expression by attenuating both p38 and c-Jun N-terminal kinase (JNK) signaling pathways. Our findings demonstrate that atRAL induces an autocrine/paracrine IL-1/IL-1R signaling to promote complement alternative pathway activation in RPE cells and provide a novel perspective on the pathomechanism of macular degeneration.

Serine protease inhibitor MDSPI16 ameliorates LPS-induced acute lung injury through its anti-inflammatory activity

A previous study described a novel serine protease inhibitor 16 from Musca domestica (MDSPI16), which inhibited the elastase and chymotrypsin. It also exhibited a potential anti-inflammatory activity for acute lung injury (ALI), while its effects on ALI are yet to be elucidated. The present study aimed to investigate the effects and the underlying mechanisms of MDSPI16 on lipopolysaccharide (LPS)-challenged mice and bone marrow neutrophils. The ALI model based on the results of LPS-induced mice demonstrated that MDSPI16 markedly reduced the infiltration of inflammatory cells, protein exudation in lung tissues, and downregulated the level of interleukin-6 (IL-6), IL-1β and tumor necrosis factor-α (TNF-α). Furthermore, the LPS-stimulated mouse bone marrow neutrophils model was employed to determine the role of MDSPI16. The cytokine levels were quantified by both the enzyme-linked immunosorbent assay (ELISA) and quantitative real-time polymerase chain reaction (qRT-PCR). Consequently, the expression of IL-6, IL-1β, and TNF-α was found to be inhibited by MDSPI16 in a dose-dependent manner. Moreover, MDSPI16 also inhibited the mouse neutrophils nuclear factor-κB (NF-κB) signaling pathway, c-Jun N-terminal kinase (JNK) signaling pathway, ERK1/2 and AP-1 signaling pathway in addition to the expression of iNOS and COX-2 proteins, which in turn, might alleviate the release of pro-inflammatory cytokines during ALI. Therefore, MDSPI16 could be proposed as a potential and novel drug therapy for ALI.

Structural basis for producing selective MAP2K7 inhibitors

Mitogen-activated protein kinase kinase 7 (MAP2K7) in the c-Jun N-terminal kinase signal cascade is an attractive drug target for a variety of diseases. The selectivity of MAP2K7 inhibitors against off-target kinases is a major barrier in drug development. We report a crystal structure of MAP2K7 complexed with a potent covalent inhibitor bearing an acrylamide moiety as an electrophile, which discloses a structural basis for producing selective and potent MAP2K7 inhibitors.

Heat shock protein 70 positively regulates transforming growth factor-α-induced hepatocellular carcinoma cell migration via the AKT signaling pathway

Heat shock proteins (HSPs) are induced in response to extracellular stress and manage the quality of proteins as molecular chaperones. HSP70, a highly conserved HSP, has been reported to correlate with the proliferation and migration of human cancer cells, such as oral, prostate, lung and liver cancer. Regarding hepatocellular carcinoma (HCC), the HSP70 levels in the tumor tissues from patients are significantly higher than those in the normal liver tissues. HSP70 reportedly upregulates the migration and invasion of HCC. The AKT, p38 mitogen-activated protein kinase (MAPK), c-jun N-terminal kinase (JNK) and Rho-kinase signaling pathways regulate the transforming growth factor (TGF)-α-induced migration of human HCC-derived HuH7 cells. However, the exact mechanism underlying the role of HSP70 in growth factor-induced HCC migration remains unclear. Therefore, in the present study, the mechanism underlying the involvement of HSP70 in TGF-α-induced HCC cell migration was investigated. Treatment with the HSP70 inhibitors VER155008 and YM-08 and the downregulation of HSP70 protein were confirmed to significantly suppress the TGF-α-induced cell migration of HuH7 cells. Both VER155008 and YM-08 reduced the TGF-α-induced phosphorylation of AKT without affecting the phosphorylation of p38 MAPK, JNK or Rho-kinase. These results strongly suggest that HSP70 positively regulates the TGF-α-induced migration of HCC cells via the AKT signaling pathway.

Peroxiredoxin 6 Attenuates Alloxan-Induced Type 1 Diabetes Mellitus in Mice and Cytokine-Induced Cytotoxicity in RIN-m5F Beta Cells.

Type 1 diabetes is associated with the destruction of pancreatic beta cells, which is mediated via an autoimmune mechanism and consequent inflammatory processes. In this article, we describe a beneficial effect of peroxiredoxin 6 (PRDX6) in a type 1 diabetes mouse model. The main idea of this study was based on the well-known data that oxidative stress plays an important role in pathogenesis of diabetes and its associated complications. We hypothesised that PRDX6, which is well known for its various biological functions, including antioxidant activity, may provide an antidiabetic effect. It was shown that PRDX6 prevented hyperglycemia, lowered the mortality rate, restored the plasma cytokine profile, reversed the splenic cell apoptosis, and reduced the β cell destruction in Langerhans islets in mice with a severe form of alloxan-induced diabetes. In addition, PRDX6 protected rat insulinoma RIN-m5F β cells, cultured with TNF-α and IL-1β, against the cytokine-induced cytotoxicity and reduced the apoptotic cell death and production of ROS. Signal transduction studies showed that PRDX6 prevented the activation of NF-κB and c-Jun N-terminal kinase signaling cascades in RIN-m5F β cells cultured with cytokines. In conclusion, there is a prospect for therapeutic application of PRDX6 to delay or even prevent β cell apoptosis in type 1 diabetes.

Lycopene ameliorates chronic stress-induced hippocampal injury and subsequent learning and memory dysfunction through inhibiting ROS/JNK signaling pathway in rats

The natural carotenoid lycopene (LYC) has strong antioxidant and neuroprotective capacities. This study investigated the effects and mechanisms of LYC on chronic stress-induced hippocampal lesions and learning and memory dysfunction. Rats were administered LYC and/or chronic restraint stress (CRS) for 21 days. Morris water maze results demonstrated that LYC prevented CRS-induced learning and memory dysfunction. Histopathological staining and transmission electron microscopy observation revealed that LYC ameliorated CRS-induced hippocampal microstructural and ultrastructural damage. Furthermore, LYC alleviated CRS-induced oxidative stress by reducing reactive oxygen species (ROS) production and enhancing antioxidant enzyme activities. LYC also improved CRS-induced hippocampal mitochondrial dysfunction by recovering mitochondrial membrane potential, and complex I (NADH dehydrogenase) and II (succinate dehydrogenase) activities. Moreover, LYC reduced CRS-induced apoptosis via the mitochondrial apoptotic pathway, and decreased the number of terminal deoxynucleotidyl transferase dUTP nick-end-labeled positive cells. Additionally, western blot analysis demonstrated that LYC inhibited CRS-induced activation of the c-Jun N-terminal kinase (JNK) signaling pathway. Correlation analysis indicated that ROS levels, JNK activation, and the mitochondrial apoptotic pathway were positively correlated. Further investigation of the underlying mechanisms revealed that the ROS scavenger N-acetyl-l-cysteine inhibited CRS-induced JNK activation. Furthermore, the JNK inhibitor SP600125 relieved CRS-induced hippocampal mitochondrial dysfunction, apoptosis via the mitochondrial apoptotic pathway, and learning and memory dysfunction. Together, these results suggest that LYC alleviates hippocampal oxidative stress, mitochondrial dysfunction, and apoptosis by inhibiting the ROS/JNK signaling pathway, thereby improving CRS-induced hippocampal injury and learning and memory dysfunction. This study provides a theoretical basis and new therapeutic strategies for the application of LYC to relieve chronic stress encephalopathy.

Abstract 4223: Combination therapy of BCR-ABL-positive B cell acute lymphoblastic leukemia by tyrosine kinase inhibitor dasatinib and c-Jun N-terminal kinase inhibition

Abstract The Philadelphia chromosome (Ph), which leads to creation and expression of the fusion gene product BCR-ABL, underlines the pathogenesis of chronic myelogenous leukemia (CML) and a fraction of adult and pediatric acute B-lymphoblastic leukemia (B-ALL). The BCR-ABL tyrosine kinase inhibitors (TKIs) has shown a remarkable clinical activity in patients with CML, but their efficacy in treating Ph+ B-ALL is limited. Identifying additional therapeutic targets is important for the effective treatment of Ph+ B-ALL. In this study, we found that the c-Jun N-terminal kinase (JNK) signaling pathway is abnormally activated in both human and mouse BCR-ABL+ B-ALL cells, but the BCR-ABL TKI does not inhibit JNK activation in these cells. Inhibition of JNK, either by RNAi-mediated downregulation or by JNK inhibitors, could significantly reduce viability of Ph+ B-ALL cells. JNK inhibition by RNAi-mediated downregulation or JNK inhibitors also showed a synergistic effect with the BCR-ABL TKI, dasatinib, in killing Ph+ B-ALL cells in vitro. Furthermore, a potent JNK inhibitor, JNK-IN-8, in combination with dasatinib markedly improved the survival of mice with BCR-ABL induced B-ALL, as compared to the treatment with dasatinib alone. Our findings indicate that simultaneously targeting both BCR-ABL and JNK kinase might serve as a promising therapeutic strategy for Ph+ B-ALL. Citation Format: Xinhua Xiao, Bo Jiao, Ruibao Ren. Combination therapy of BCR-ABL-positive B cell acute lymphoblastic leukemia by tyrosine kinase inhibitor dasatinib and c-Jun N-terminal kinase inhibition [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 4223.

MiR-128 regulated the proliferation and autophagy in porcine adipose-derived stem cells through targeting the JNK signaling pathway

Purpose microRNA-128 (miR-128), a brain-enriched microRNA, has been reported to play a crucial role in the treatment of diseases. The c-Jun N-terminal kinase (JNK) signaling pathway exerts various biological functions such as regulation of cell proliferation, differentiation and apoptosis. In this study, we investigated the role of the miRNA-128-JNK signaling pathway in proliferation, apoptosis and autophagy of porcine adipose-derived stem cells (ASCs). Methods After over-expressing miR-128 in porcine ASCs, cell proliferation was determined by 2,3-Bis-(2-Methoxy-4-Nitro-5-Sulfophenyl)-2H-Tetrazolium-5-Carboxanilide (XTT) method, cell apoptosis was observed by Flow cytometry (FCM), the expression of miR-128, B-cell lymphoma 2 (Bcl-2), and Bcl-2-associated X protein (Bax) was measured by RNA preparation and reverse transcription polymerase chain reaction (RT-PCR), and protein expression of JNK, phosphorylated JNK (p-JNK) and LC3B was analyzed by Western Blot analysis. Results The over-expression of miR-128 potently promoted cell proliferation and autophagy while suppressed the apoptosis of porcine ASCs. In addition, the down-regulated expression level of p-JNK was detected in miR-128-over-expressed porcine ASCs. However, followed by the block of the JNK signaling pathway using SP600125 inhibitor, the effects of miR-128 on the proliferation, apoptosis and autophagy of porcine ASCs were significantly suppressed. Conclusion It is demonstrated that the miR-128-JNK signaling pathway is a potential therapeutic target for the treatment of obesity.